The objective of the proposed research project is to test the hypothesis that weight loss, improved glucose homeostasis, and increased energy expenditure following Roux-en-Y gastric bypass (RYGB) surgery are, in part, mediated by a change in the gut microbial community. RYGB is currently the most effective long-term surgical treatment option for both diabetes and obesity, resulting in vast improvements in insulin sensitivity prior to, and independent of, marked and sustained weight loss. Physical manipulation of the gut results in an altered flow of nutrients and gastrointestinal secretions, which likely affects gut-derived signaling pathways regulating energy balance and glucose metabolism. The microorganisms that reside in the gastrointestinal tract (the gut microbiota) are heavily influenced by diet and have recently been implicated as powerful regulators of obesity-associated fat deposition, decreased energy expenditure, and insulin resistance. Human and rat studies have demonstrated that the gut microbial ecology changes markedly following RYGB and is different from the microbial communities observed in either lean or obese individuals. Given these observed shifts in this ecology after surgery, and given the emerging role of the microbiota in regulating energy balance and glucose metabolism, the gut microbiota is a likely mediator of the effects of RYGB. However, the extent to which changes in gut microbial ecology contribute to post-RYGB metabolic outcomes remains unclear. The proposed studies aim to characterize the degree to which the gut microbial community is changed after RYGB in the mouse and to what degree these changes mediate RYGB outcomes. Gut microbial changes will be assessed by 16S rRNA gene sequencing of fecal samples, collected longitudinally over time after surgery, and of luminal and mucosal adherent communities collected along the gastrointestinal axis. To determine the extent to which these microbial changes mediate metabolic outcomes, germ free mice will be inoculated with gut contents from donor animals that had undergone either RYGB or SHAM treatment. These recipient animals will then be assessed for changes in body weight, adiposity, food intake, energy expenditure, and glucose parameters. Subsequently, we will determine which metabolic signaling pathways are most likely impacted by colonization of microbiota from RYGB donors in the gnotobiotic animals, based on the physiologic phenotypes that are observed. Successful completion of the proposed project will increase our understanding of the physiological role of the gastrointestinal tract in regulating metabolic function, including the contribution of the microbiota on afferent signaling i the gut, as well as enable identification of new therapeutic targets to treat obesity and diabetes that will mimic the physiology of RYGB without the need for surgery itself. PUBLIC HEALTH RELEVANCE: This project aims to understand how the trillions of micro-organisms that live in the gut are affected by Roux- en-Y gastric bypass, a surgical intervention that has profound beneficial effects on energy balance, glucose homeostasis, and other metabolic functions, and to what extent changes in gut bacterial populations affect these metabolic improvements. The proposed studies will include assessments of gut microbial contributions to multiple aspects of metabolic regulation on both a physiological and molecular level, including food intake, fat deposition and body composition, energy expenditure, lipid metabolism and glucose metabolism.